猫Darkschewitsch氏核向大脑皮质的投射——HRP逆轴突传递法研究

本文用HRP逆行传递法研究了猫中脑Darkschewitsch氏核(DK)向大脑皮质的投射,观察了起源细胞的形态和分布。实验证明DK仅投向前、后乙状回(感觉运动皮质),其余各脑回或机能区均不接受DK的投射;DK只投向感觉运动皮质的前肢区,而不投向后肢区和面区。标记细胞以中等大小的卵圆形、梭形和三角形为主,散在分布于同侧DK的全长,对侧几乎无标记。核的吻段较尾段的标记细胞多。

Potentilla anserina polysaccharide alleviates cadmium-induced oxidative stress and apoptosis of H9c2 cells by regulating the MG53-mediated RISK pathway

Oxidative stress plays a crucial role in cadmium(Cd)-induced myocardial injury.Mitsugumin 53(MG53)and its mediated reperfusion injury salvage kinase(RISK)pathway have been demonstrated to be closely related to myocardial oxidative damage.Potentilla anserina L.polysaccharide(PAP)is a polysaccharide with antioxidant capacity,which exerts protective effect on Cd-induced damage.However,it remains unknown whether PAP can prevent and treat Cd-induced cardiomyocyte damages.The present study was desgined to explore the effect of PAP on Cd-induced damage in H9c2 cells based on MG53 and the mediated RISK pathway.For in vitro evaluation,cell viability and apoptosis rate were analyzed by CCK-8 assay and flow cytometry,respectively.Furthermore,oxidative stress was assessed by 2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA)staining and using superoxide dismutase(SOD),catalase(CAT),and glutathione/oxidized glutathione(GSH/GSSG)kits.The mitochondrial function was measured by JC-10 staining and ATP detection assay.Western blot was performed to detect the expression of proteins related to MG53,the RISK pathway,and apoptosis.The results indicated that Cd increased the levels of reactive oxygen species(ROS)in H9c2 cells.Cd decreased the activities of SOD and CAT and the ratio of GSH/GSSG,resulting in decreases in cell viability and increases in apoptosis.Interestingly,PAP reversed Cd-induced oxidative stress and cell apoptosis.Meanwhile,Cd reduced the expression of MG53 in H9c2 clls and inhibited the RISK pathway,which was mediated by decreasing the ratio of p-Akt^(Ser473)/Akt,p-GSK3β^(Ser9)/GSK3β and p ERK1/2/ERK1/2.In addition,Cd impaired mitochondrial function,which involved a reduction in ATP content and mitochondrial membrane potential(MMP),and an increase in the ratio of Bax/Bcl-2,cytoplasmic cytochrome c/mitochondrial cytochrome c,and Cleaved-Caspase 3/Pro-Caspase 3.Importantly,PAP alleviated Cd-induced MG53 reduction,activated the RISK pathway,and reduced mitochondrial damage.Interestingly,knockdown of MG53 or inhibition of the RISK pathway attenuated the protective effect of PAP in Cd-induced H9c2 cells.In sum,PAP reduces Cd-induced damage in H9c2 cells,which is mediated by increasing MG53 expression and activating the RISK pathway.

Protective effect of resveratrol on rat cardiomyocyte H9C2 cells injured by hypoxia/reoxygenation by regulating mitochondrial autophagy via PTEN-induced putative kinase protein 1/Parkinson disease protein 2 signaling pathway

OBJECTIVE:To investigate the protective effect of resveratrol on cardiomyocytes after hypoxia/reoxygenation intervention based on PTEN-induced putative kinase protein 1/Parkinson disease protein 2(PINK1/PARKIN)signaling pathway.METHODS:3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide was used to detect the effect of resveratrol on the viability of H9C2 cells;the hypoxia/reoxygenation(H/R)model was established in tri-gas incubator;2’,7’-Dichlorofluorescin diacetate staining was used to measure the content of reactive oxygen species(ROS);the changes of mitochondrial membrane potential was determined by 5,5’,6,6’-Tetrachloro-1,1’,3,3’-tetraethyl-imidacarbocyanine iodide staining;the changes of mitochondrial respiratory chain complex activity was evaluated by enzyme activity kits;flow cytometry was used to detect the ratio of apoptotic cells;transmission electron microscope was used to observe the ultrastructure of H9C2 cells;Western blot was used to detect the protein changes of mitochondrial 20 k Da outer membrane protein(TOM20),translocase of inner mitochondrial membrane 23(TIM23),presenilins associated rhomboid-like protein(PARL),PINK1,PARKIN and mitofusin 1(Mfn1),mitofusin 2(Mfn2),phosphotyrosine independent ligand for the Lck SH2 domain of 62 k Da(P62),microtubule-associated protein 1 light chain 3 beta(LC3B);the m RNA levels of PINK1 and PARKIN was detected by quantitative polymerase chain reaction;immunoprecipitation assay was used to detect the interaction between PARKIN and Ubiquitin.RESULTS:Resveratrol could inhibit the proliferation of H9C2 cells in a time-and concentration-dependent manner;however,pretreatment with low cytotoxic resveratrol could reduce the H/R-induced increase in cellular ROS levels,alleviate the loss of mitochondrial membrane potential induced by H/R,inhibit H/R-induced apoptosis of H9C2 cells,and protect the mitochondrial structure and respiratory chain of H9C2 cells from H/R damage.Resveratrol could further increase the levels of p62,PINK1,PARKIN protein,the expression of PINK1,PARKIN m RNA and the ratio of LC3BⅡ/LC3BⅠin H/Rinduced H9C2 cells,inhibit the interaction between PARKIN and Ubiquitin in H/R-induced H9C2 cells,and further reduce the expression of TOM20,TIM23,PARL,Mfn1 and Mfn2 protein in H/R-induced H9C2 cells.The effect of resveratrol is consistent with that of autophagy activator on H/R-induced H9C2 cells.CONCLUSIONS:Resveratrol can protect H9C2 cells from H/R injury,which may be related to resveratrol promoting mitochondrial autophagy by activating PINK1/PARKIN signaling pathway.